PROJECT SUMMARY The overall objective of this proposal is to understand how dietary energy balance and especially obesity influences genome stability at endogenous mutation ?hotspots?. To achieve this objective, we will use novel mouse models that we have developed to determine the impact of obesity on DNA structure-induced mutagenesis in various tissues from these mice. Repetitive DNA sequences are widely dispersed throughout mammalian genomes and can adopt alternative (non-B DNA) secondary structures, such as H-DNA and Z- DNA. Importantly, these non-B DNA structure-forming sequences are significantly enriched at endogenous mutation hotspots in human cancer genomes, implicating them in cancer etiology. For example, H-DNA- forming sequences in the c-MYC gene are found at translocation breakage hotspots in Burkitt?s lymphoma and acute B-cell lymphoma. We have developed novel mutation-reporter mice containing human H-DNA- and Z- DNA-forming sequences that co-localize with translocation breakpoint hotspots, and demonstrated for the first time that these sequences are mutagenic in vivo. Obesity is known to increase cellular oxidative stress and oxidative DNA damage, whereas calorie restriction has been shown to decrease cellular oxidative stress, oxidative DNA damage, reduce mutagenesis and to enhance DNA repair pathways. In addition, obesity is an important risk factor for a significant number of cancers. However, the extent to which dietary energy balance and especially obesity influences DNA structure-induced genetic instability is not known. Thus, a goal of the proposed work is to fill this gap in knowledge. In this proposal, we will test the working hypothesis that obesity increases DNA structure-induced genetic instability. We will examine the impact of diet-induced obesity and calorie restriction on DNA structure-induced mutagenesis in mice. Several different tissues will be evaluated from these mice to determine whether there are any tissue specific differences in mutagenesis in response to energy balance manipulation. We will also explore mechanisms for the impact of obesity on DNA structure- induced genetic instability. We will focus our studies on the impact of obesity on DNA repair mechanisms as several recent studies have suggested that obesity impairs multiple DNA repair pathways, including non- homologous end-joining, a repair pathway that we have found to play a role in the processing of non-B DNA. In addition, we have found that flap endonuclease 1 (FEN1) plays an important role in error-free processing of non-B DNA and that its levels are modulated by dietary energy balance. These observations will also be explored in more detail in this proposal. Completion of the proposed studies will lead to a greater understanding of how obesity influences cancer etiology. In addition, this work will lead to the identification of novel targets for the prevention and/or treatment of obesity-related cancers.